T CELL HYBRIDOMA LYMPHOKINE CHEMOTACTIC FOR T LYMPHOCYTE

局部T淋巴细胞聚集被认为是肿瘤的重要因素 并可能预示着局部自身免疫性疾病和排斥反应 移植的器官。高血压病患者T淋巴细胞聚集机制的研究 最初抗原刺激的部位没有很好的定义，但它是已知的 抗原刺激的T细胞产生一种淋巴因子(S)。 对其他T细胞的趋化作用。 据认为，几种不同的淋巴因子可以特异性地吸引 辅助性/诱导性或抑制性/细胞毒性T细胞可在 不同的时间，可能是通过不同的T淋巴细胞亚群 在局部炎症反应的整个过程中。更进一步说，它是 这些淋巴因子可能在决定 在局部炎症反应中聚集的淋巴细胞亚群 从而调节免疫反应。 一种能产生大量T细胞的T细胞杂交瘤FS730.11 特异性淋巴细胞趋化因子(LCF)，但仅限于有限范围 其他淋巴因子已被开发出来。Con A刺激的上清来自 FS730.11的培养将作为单个细胞的低密度脂蛋白的来源 类型以定义辅助性或抑制性T细胞亚群是否响应 到这个特殊的LCF。辅助性和抑制性T细胞亚群将 将尼龙毛非贴壁小鼠脾细胞接种于大鼠体内获得 抗Lyt1或Lyt2单抗包被塑料组织培养 车牌。细胞亚群的纯度将通过Flow进行评估 细胞计量学。这些T细胞亚群的趋化性，作为对LCF的响应， 然后通过8微米孔径的迁移进行评估 改进型博伊登盲室中的硝化棉过滤器。 将半纯化的LCF注射到小鼠的腹膜内。这个 苏木素对腹膜渗出细胞进行量化、分化 免疫组化染色，流式细胞仪检测T淋巴细胞。 低碳纤维的提纯将通过顺序超滤和 高效液相色谱分析。LCF将被描述为 通过对几种酶的失活敏感，通过圆盘凝胶 电泳法、等电聚焦法和柱层析法。 纯化的小鼠LCF将在大鼠体内产生抗体。从本质上讲， 放射性标记的LCF将由ConA刺激FS730.11产生 组织培养，建立了LCF的放射免疫分析(RIA)方法。此RIA 将为未来实验确定T细胞亚群提供便利(S) 低碳纤维的来源和低碳纤维的生产动力学。一株大鼠B细胞杂交瘤 将产生分泌抗LCF抗体，来自B细胞的抗体 细胞杂交瘤将在未来的实验中用于研究 体内通过抑制LCF活性的免疫反应。

Local T lymphocyte accumulation is thought to be important for tumor rejection and may presage local autoimmune disease and rejection of transplanted organs. The mechanisms of T lymphocyte accumultion at the site of initial antigen stimulation are not well defined, but it is known that antigen stimulated T cells produce a lymphokine(s) that is (are) chemotactic for other T cells. It is proposed that several different lymphokines that specifically attract either helper/inducer or suppressor/cytotoxic T cells may be released at different times and perhaps by different T lymphocyte subpopulations throughout the course of a local inflammatory response. Further, it is probable that these lymphokines play a significant role in determining the lymphocyte subpopulations that accumulate in a local inflammatory response and thereby, modulate the immune response. A T cell hybridoma, FS730.11, that produces large amounts of T cell specific lymphocyte chemotactic factor (LCF), but only a restricted range of other lymphokines has been developed. Con A stimulated supernate from cultures of FS730.11 will be used as a source of LCF from a single cell type to define whether helper or suppressor T cell subpopulations respond to this particular LCF. Helper and suppressor T cell subpopulations will be obtained by plating nylon wool non-adherent, mouse spleen cells on rat monoclonal anti-Lyt1 or Lyt2 antibody-coated plastic tissue culture plates. The purity of cell subpopulations will be assessed by flow cytrometry. Chemotaxis of these T cell subpopulations, in response to LCF, will then be assessed by migration through 8 micron pore size nitrocellulose filters in modified blindwell Boyden chambers. Semi-purified LCF will be injected into the peritoneum of mice. The peritoneal exudate cells will be quantified, differentiated by hematoxylin staining, and evaluated by flow cytometry for T lymphocyte. Purification of LCF will be initiated by sequential ultrafiltration and high performance liquid chromatography (HPLC). LCF will be characterized by sensitivity to inactivation with several enzymes, by disc gel electrophoresis, by isoelectric focusing and by column chromatography. Antibody to purified mouse LCF will be produced in rats. Intrinsically, radiolabelled LCF will be produced by Con A stimulation of FS730.11 in tissue culture, and a radioimmunoassay (RIA) for LCF developed. This RIA will facilitate future experiments to determine the T cell subpopulation(s) source of LCF and the kinetics of LCF production. A rat B cell hybridoma that secretes anti-LCF antibody will be produced, and antibody from that B cell hybridoma will be used in future experiments to study modulation of the immune response by inhibiting LCF activity in vivo.